The genetic redirection of T cells with chimeric antigen receptors (CARs) that link an antigen-specific single-chain antibody fragment (scFv) to intracellular signaling domains is at the forefront of cancer immunotherapy (Jena et al., 2010, Blood 116:1035-1044; Gross et al., 1989, Proc. Natl. Acad. Sci. USA 86:10024-10028; Porter et al., 2011, N. Engl. J. Med. 365:725-733). CARs can functionally redirect T cells with high specificity to various surface antigens on tumor cells independent of MHC restriction and antigen processing, and therefore bypass major mechanisms by which tumors escape immune recognition. T cells bearing a first generation CAR having only the T cell CD3z intracellular signaling domain either fail to persist or become anergic since tumor cells frequently lack appropriate ligands for costimulatory molecules (Inman et al., 2007, Curr. Cancer Drug Targets 7:15-30). Indeed, incomplete activation of CART cells in vivo appears to limit their expansion and persistence in vivo, and thus hampered their efficacy in clinical trials in subjects with lymphoma (Till et al., 2008, Blood 112:2261-2271), neuroblastoma (Pule et al., 2008, Nat. Med. 14:1264-1270), or ovarian (Kershaw et al., 2006, Clin. Cancer Res. 12:6106-6115) or renal cancer (Lamers et al., 2006, J. Clin. Oncol. 24:e20-22).
To overcome these limitations, second generation CART cells were developed that incorporate the intracellular domain of various costimulatory molecules such as CD28, 4-1BB, OX-40, and CD27, leading to superior expansion, persistence and activity of the CART cells in preclinical mouse models (Carpenito et al., 2009, Proc. Natl. Acad. Sci. USA 106:3360-3365; Song et al., 2012, Blood 119:696-706) and in clinical studies (Porter et al., 2011, N. Engl. J. Med. 365:725-733; Kalos et al., 2011, Sci. Transl. Med. 3:95ra73; Savoldo et al., 2011, J. Clin. Invest. 121:1822-1826). However, the enhanced potency of the CARs can be associated with autoimmunity due to on-target toxicities against normal tissues expressing the lower levels of the tumor associated antigens, with two serious adverse events (SAEs) reported so far.
The first SAE occurred in a clinical trial where administration of anti-ErbB2 CART cells containing the CD28 and 4-1BB costimulatory signaling regions into a patient with refractory colon cancer with metastatic sites in lung and liver led to development of dramatic pulmonary toxicity with lung infiltrates and a cytokine storm followed by cardiac arrest and patient death (Morgan et al., 2010, Cancer J. 16:336-341). In the second report, a lymphodepleted patient with bulky chronic lymphocytic leukemia received autologous T cells engineered with an anti-CD19 second-generation CAR containing the CD28 domain at a total dose of 3×107 T cells/kg. This patient developed fever, hypotension, and dyspnea 20 hours after infusion, which rapidly progressed. Low grade sepsis was the most likely reason of the patient's death, however the possibility that a cyclophosphamide-induced “cytokine storm” may have enhanced the in vivo activation of modified T cells is well considered (Brentjens et al., 2010, Mol. Ther. 18:666-668). It is therefore clear that the development of strategies limiting potential early or late toxicity is mandatory.
A fully human anti-mesothelin CAR capable of conferring potent in vitro and in vivo effector functions to primary T cells against mesothelin-expres sing tumors has previously been generated (Lanitis et al., 2012, Mol. Ther. 20:633-643). However, mesothelin CART cells hold the potential to inflict damage against normal mesothelial cells lining the pleura, peritoneum and peritoneum as well as epithelial cells of the trachea, tonsils, fallopian tube and the rete testis which express low levels of mesothelin (Chang et al., 1992, Int. J. Cancer 50:373-381; Ordonez, 2003, Mod. Pathol. 16:192-197).
There is thus a need in the art for CAR therapies that do not exhibit serious adverse events. The present invention addresses this unmet need in the art.